Deletion of the Mycobacterium tuberculosis α-crystallin-like hspX gene causes increased bacterial growth in vivo

Hypervirulent mutants of Mycobacterium tuberculosis, whose growth rates are higher in vivo, have now been reported to have mutations in both regulatory and structural genes, but the basis for this unusual phenotype is not understood. One hypervirulence gene, dosR (devR, Rv2031c), activates transcription of approximately 50 genes in this pathogen in response to hypoxia and nitric oxide stress. The most dramatic activation (similar to 80-fold) is activation of the hspX (acr, Rv2031c) gene, which encodes a 16-kDa alpha-crystallin-like protein that is a major antigen. In this study we found that a Delta acr mutant exhibited increased growth following infection of BALB/c mice in vivo and in both resting and activated macrophages in vitro (as measured by the number of CFU). The increased growth in macrophages was equal to that of a Delta dosR mutant, while introduction of a constitutively expressed hspX gene reduced the Delta dosR virulence to wild-type levels. These results suggest that the increased number of CFU of the Delta dosR mutant was largely due to loss of hspX expression. We also confirmed that constitutive expression of hspX slows growth in vitro, and we propose that hspX plays an active role in slowing the growth of M. tuberculosis in vivo immediately following infection.